RRC ID 69995
Author Inoue M, Takeuchi A, Horigane S, Ohkura M, Gengyo-Ando K, Fujii H, Kamijo S, Takemoto-Kimura S, Kano M, Nakai J, Kitamura K, Bito H.
Title Rational design of a high-affinity, fast, red calcium indicator R-CaMP2.
Journal Nat Methods
Abstract Fluorescent Ca(2+) reporters are widely used as readouts of neuronal activities. Here we designed R-CaMP2, a high-affinity red genetically encoded calcium indicator (GECI) with a Hill coefficient near 1. Use of the calmodulin-binding sequence of CaMKK-α and CaMKK-β in lieu of an M13 sequence resulted in threefold faster rise and decay times of Ca(2+) transients than R-CaMP1.07. These features allowed resolving single action potentials (APs) and recording fast AP trains up to 20-40 Hz in cortical slices. Somatic and synaptic activities of a cortical neuronal ensemble in vivo were imaged with similar efficacy as with previously reported sensitive green GECIs. Combining green and red GECIs, we successfully achieved dual-color monitoring of neuronal activities of distinct cell types, both in the mouse cortex and in freely moving Caenorhabditis elegans. Dual imaging using R-CaMP2 and green GECIs provides a powerful means to interrogate orthogonal and hierarchical neuronal ensembles in vivo.
Volume 12(1)
Pages 64-70
Published 2015-1-1
DOI 10.1038/nmeth.3185
PII nmeth.3185
PMID 25419959
MeSH Action Potentials / physiology Animals Caenorhabditis elegans / radiation effects Calcium / metabolism Calcium Signaling / physiology Calcium-Calmodulin-Dependent Protein Kinase Kinase / metabolism* Calmodulin-Binding Proteins Cells, Cultured Cerebral Cortex / cytology Fluorescent Dyes / metabolism HEK293 Cells Hippocampus / cytology Humans Indicators and Reagents / chemical synthesis* Light Mice Neurons / physiology Patch-Clamp Techniques Peptide Fragments / chemistry Peptide Fragments / metabolism
IF 30.822
DNA material pBS-CaMKIIpro-R-CaMP2-WPRE (RDB19475)